Haiqing Chen ¹ Haotian Lai ² Hao Chi ² Wei Fan ^2,3,4 Jinbang Huang ² Shengke Zhang ² Chenglu Jiang ² Lai Jiang ² Qingwen Hu ² Xiuben Yan ^2* Yemeng Chen ^5* Jieying Zhang ^6* Guanhu Yang ^7* Bin Liao ^2,3,4* Juyi Wan ^2,3,4*

• ¹ Southwest Medical University, Luzhou, China
• ² School of Clinical Medicine, The Affiliated hospital, Southwest Medical University, Luzhou, China, Luzhou, China
• ³ Department of Cardiovascular Surgery, The Affiliated Hospital, Southwest Medical University, Luzhou, China
• ⁴ Key Laboratory of Medical Electrophysiology, Ministry of Education, Institute of Cardiovascular Medicine, Southwestern Medical University, Luzhou, Sichuan Province, China
• ⁵ New York College of Traditional Chinese Medicine, Mineola, New York, United States
• ⁶ First Teaching Hospital of Tianjin University of Traditional Chinese Medicine, Nankai District, Tianjin, China
• ⁷ Department of Specialty Medicine, Ohio University, Athens, Ohio, United States

Background: Atherosclerosis, a complex chronic vascular disorder with multifactorial etiology, stands as the primary culprit behind consequential cardiovascular events, imposing a substantial societal and economic burden. Nevertheless, our current understanding of its pathogenesis remains imprecise. In this investigation, our objective is to establish computational models elucidating molecular-level markers associated with atherosclerosis.This endeavor involves the integration of advanced machine learning techniques and comprehensive bioinformatics analyses.Our analysis incorporated data from three publicly available the Gene Expression Omnibus (GEO) datasets: GSE100927 (104 samples, 30,558 genes), which includes atherosclerotic lesions and control arteries from carotid, femoral, and infra-popliteal arteries of deceased organ donors; GSE43292 (64 samples, 23,307 genes), consisting of paired carotid endarterectomy samples from 32 hypertensive patients, comparing atheroma plaques and intact tissues; and GSE159677 (30,498 single cells, 33,538 genes), examining single-cell transcriptomes of calcified atherosclerotic core plaques and adjacent carotid artery tissues from patients undergoing carotid endarterectomy. Utilizing single-cell sequencing, highly variable atherosclerotic monocyte subpopulations were systematically identified. We analyzed cellular communication patterns with temporal dynamics. The bioinformatics approach Weighted Gene Co -expression Network Analysis (WGCNA) identified key modules, constructing a Protein-Protein Interaction (PPI) network from module-associated genes. Three machine-learning models derived marker genes, formulated through logistic regression and validated via convolutional neural network(CNN) modeling. Subtypes were clustered based on Gene Set Variation Analysis (GSVA) scores, validated through immunoassays.: Three pivotal atherosclerosis-associated genes-CD36, S100A10, CSNK1A1were unveiled, offering valuable clinical insights. Profiling based on these genes delineated two distinct isoforms: C2 demonstrated potent microbicidal activity, while C1 engaged in inflammation regulation, tissue repair, and immune homeostasis. Molecular docking analyses explored therapeutic potential for Estradiol, Zidovudine, Indinavir, and Dronabinol for clinical applications. Conclusion: This study introduces three signature genes for atherosclerosis, shaping a novel paradigm for investigating clinical immunological medications. It distinguishes the high biocidal C2 subtype from the inflammation-modulating C1 subtype, utilizing identified signature gene as crucial targets.